Longitudinal Assessment of the Dynamics of Escherichia coli, Total Coliforms, Enterococcus spp., and Aeromonas spp. in Alternative Irrigation Water Sources: a CONSERVE Study

Abstract

As climate change continues to stress freshwater resources, we have a pressing need to identify alternative (nontraditional) sources of microbially safe water for irrigation of fresh produce. This study is part of the center CONSERVE, which aims to facilitate the adoption of adequate agricultural water sources. A 26-month longitudinal study was conducted at 11 sites to assess the prevalence of bacteria indicating water quality, fecal contamination, and crop contamination risk (Escherichia coli, total coliforms [TC], Enterococcus, and Aeromonas). Sites included nontidal freshwater rivers/creeks (NF), a tidal brackish river (TB), irrigation ponds (PW), and reclaimed water sites (RW). Water samples were filtered for bacterial quantification. E. coli, TC, enterococci (∼86%, 98%, and 90% positive, respectively; n = 333), and Aeromonas (∼98% positive; n = 133) were widespread in water samples tested. Highest E. coli counts were in rivers, TC counts in TB, and enterococci in rivers and ponds (P < 0.001 in all cases) compared to other water types. Aeromonas counts were consistent across sites. Seasonal dynamics were detected in NF and PW samples only. E. coli counts were higher in the vegetable crop-growing (May-October) than nongrowing (November-April) season in all water types (P < 0.05). Only one RW and both PW sites met the U.S. Food Safety Modernization Act water standards. However, implementation of recommended mitigation measures of allowing time for microbial die-off between irrigation and harvest would bring all other sites into compliance within 2 days. This study provides comprehensive microbial data on alternative irrigation water and serves as an important resource for food safety planning and policy setting.IMPORTANCE Increasing demands for fresh fruit and vegetables, a variable climate affecting agricultural water availability, and microbial food safety goals are pressing the need to identify new, safe, alternative sources of irrigation water. Our study generated microbial data collected over a 2-year period from potential sources of irrigation (rivers, ponds, and reclaimed water sites). Pond water was found to comply with Food Safety Modernization Act (FSMA) microbial standards for irrigation of fruit and vegetables. Bacterial counts in reclaimed water, a resource that is not universally allowed on fresh produce in the United States, generally met microbial standards or needed minimal mitigation. We detected the most seasonality and the highest microbial loads in river water, which emerged as the water type that would require the most mitigation to be compliant with established FSMA standards. This data set represents one of the most comprehensive, longitudinal analyses of alternative irrigation water sources in the United States.

Keywords: Aeromonas; Food Safety Modernization Act; fecal indicators; food safety; irrigation water; irrigation water physicochemical parameters.

FIG 1

Bacterial prevalence in log CFU/100 ml in various water types for E. coli (A), total coliforms (TC) (B), Enterococcus spp. (C), and Aeromonas spp. (D). Data for each water type are pooled from various sites: nontidal fresh river (NF; n = 166 from 5 sites), pond water (PW; n = 69 from 2 sites), reclaimed water (RW; n = 64 from 3 sites), and tidal brackish rivers (TB; n = 34 from 1 site). The boxplots show the median and the 25th and 75th percentiles of the range. The whiskers show lower and higher observations than the 25th and 75th percentiles, respectively. Lowercase letters denote statistically significant differences at a P value of <0.05 among water types for each taxon.

FIG 2

Seasonal variation in bacterial counts in log CFU/100 ml for E. coli (A), total coliforms (TC) (B), Enterococcus spp. (C), and Aeromonas spp. (D), enumerated in different water types. Data for each water type are pooled from various sites. The boxplots show the median and the 25th and 75th percentiles of the range. The whiskers show lower and higher observations than the 25th and 75th percentiles, respectively. Lowercase letters denote statistically significant differences at a P value of <0.05 among water types for each taxon.

FIG 3

Bacterial counts in log CFU/100 ml for E. coli (A), total coliforms (TC) (B), Enterococcus spp. (C), and Aeromonas spp. (D), enumerated in different water types and categorized by vegetable crop-growing (light gray bars) and nongrowing (dark gray bars) seasons. Asterisks indicate a significant difference by Student's t test. **, P ≤ 0.01; *, P ≤ 0.05. The boxplots show the median and the 25th and 75th percentiles of the range. The whiskers show lower and higher observations than the 25th and 75th percentiles, respectively.

FIG 4

(A) Geometric means (GM) and statistical threshold values (STV) of E. coli counts in log CFU/100 ml for vegetable crop-growing season months only for 11 sites, including nontidal rivers (NF), tidal rivers (TB), pond water (PW), and reclaimed water (RW) sites, in relation to the Produce Safety Rule (PSR) standards. Light gray bars indicate GM, dark gray bars indicate STV, the light gray dashed line indicates PSR GM threshold value of 2.1 log CFU/100 ml, and the dark gray dashed line indicates PSR STV threshold value of 2.61 log CFU/100 ml. The table in the figure displays delays needed between application of irrigation water and harvest to allow for bacterial die-off, stipulated in the PSR to occur at a rate of 0.5 log CFU/day. (B to E) Month-to-month variation in average E. coli counts over two growing seasons in nontidal rivers (NF) (B), tidal brackish water (TB) (C), pond water (PW) (D), and reclaimed water (RW) (E). Different lowercase letters denote statistical differences at a P value of <0.05 in bacterial counts by month of collection, and error bars denote standard errors.

FIG 5

Relationships between bacterial indicator counts for E. coli, total coliforms (TC), and Enterococcus spp. in log CFU/100 ml by season (A to C) and water type (D to F). R² values indicate goodness of fit of the line using Pearson correlation analysis. NF denotes nontidal river, PW denotes pond water, RW denotes reclaimed water, and TB denotes tidal river.

FIG 6

Relationships between Aeromonas species counts and bacterial indicator counts (E. coli, total coliforms [TC], and Enterococcus spp.) in log CFU/100 ml by season (A to C) and water type (D to F). R² values indicate goodness of fit of the line using Pearson correlation analysis and are given in each panel. NF denotes nontidal river, PW denotes pond water, RW denotes reclaimed water, and TB denotes tidal river.

FIG 7

Relationship between bacterial counts and physicochemical parameters in different water types. EC indicates E. coli, TC indicates total coliforms, Ent indicates Enterococcus spp., and Aer indicates Aeromonas spp. Values in each box indicate Pearson correlation coefficient (r) at a P value of <0.05. Pink indicates positive and blue indicates negative association between two variables (bacterial counts and physicochemical parameter). The intensity of the box color increases with increasing value of r.